1. Field of the Invention
The present invention generally relates to methods and compositions for screening for antiviral compounds using cell cultures. More specifically, the invention relates to the use of multiple cell cultures harboring multiple subgenomic viral replication systems to evaluate compounds for antiviral activity.
2. Description of the Related Art
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Primary screening programs to discover and identify compounds with antiviral activity can be designed in a variety of ways. All programs, however, fall into one of two general approaches. In the targeted approach, one particular biochemical target is chosen and candidate antiviral compounds are screened for inhibition of that target. The target is often an enzyme or a receptor that is known or thought to be essential to the process of viral replication. The alternative approach is unbiased such that inhibitors of viral replication are sought without a priori concern for the target. This unbiased approach generally involves use of cell culture since, as obligate intracellular pathogens, viruses can only replicate within cells. Although cell-based screening has been used successfully throughout the drug-discovery field, it is problematic when screening for antivirals. This is because it requires inoculation of infectious virus onto the cells and the production of additional infectious progeny virus. In particular, handling such infectious material is not easily compatible with the high throughput process of screening large libraries of compounds.
Thus, there is a need for improved methods and compositions that are useful for screening and analyzing antiviral compounds. In particular, these methods and compositions should be useful for high-throughput antiviral screening. The invention described herein satisfies that need.
Accordingly, the present invention provides methods and compositions utilizing subgenomic viral replication systems to evaluate potential antiviral compounds. The use of these methods and compositions allows rapid screening of the potential antiviral compounds against multiple viruses simultaneously.
In some embodiments, the present invention is directed to methods of screening a candidate antiviral agent for antiviral activity. The methods comprise the use of at least two subgenomic viral replication systems that are genetically distinct from each other. The methods involve preparing a first cell culture comprising cells containing a first subgenomic viral replication system, and a second cell culture comprising cells containing a second subgenomic viral replication system, adding the candidate antiviral agent to each cell culture, incubating the cell cultures under conditions and for a time sufficient to detect an antiviral effect by the candidate antiviral agent on the subgenomic viral replication systems, and determining the effect of the candidate antiviral agent on each viral replication system.
Related embodiments of the present invention are directed to methods of screening a candidate antiviral agent for antiviral activity. These methods also comprise the use of at least two subgenomic viral replication systems that are genetically distinct from each other. These methods include combining a first cell culture comprising cells containing a first subgenomic viral replication system and a second cell culture comprising cells containing a second subgenomic viral replication system to make a mixed cell culture, adding the candidate antiviral agent to the mixed cell culture, incubating the mixed cell culture under conditions and for a time sufficient to detect an antiviral effect by the candidate antiviral agent on the subgenomic viral replication systems, and determining the effect of the candidate antiviral agent on each viral replication system.
The present invention is also directed to mixed cell cultures that include a first cell culture comprising cells containing a first subgenomic viral replication system and a second cell culture comprising cells containing a second subgenomic viral replication system.